1. Field of the Invention
This invention relates to the diagnosis and monitoring of ischemia, including but not limited to myocardial and cerebral ischemia, by measuring the concentration of molecules that do not originate from the ischemic tissue but whose concentration in the blood and other fluids changes as a consequence of the ischemic state.
2. Description of the Related Art
Ischemia is a reduction in blood flow. This reduction may occur for a variety of reasons, including but not limited to thrombosis, embolism, aneurysm, spasm, or collapse of a blood vessel due to deterioration. Because of the reduction in blood flow the tissue that would otherwise be nourished may no longer receive sufficient nutrition to maintain cellular integrity, it also may not be able to remove sufficient amounts of cellular waste products and it may also result in inadequate exchange of blood gases such as oxygen and carbon dioxide. The inability to transfer sufficient oxygen to the cells (hypoxia) may have many of the same consequences of ischemia and can also be detected by the same methods claimed in this invention.
If ischemia persists for a sufficient time, that is, if oxygenated blood flow is not restored, then the cells of the tissue normally perfused by the blood flow, will begin to die. This may occur gradually over time and may be unnoticed until sufficient cell destruction has occurred so that the function of the organism is significantly impaired. An example may be the gradual deterioration of circulation to the extremities or other body parts that occurs in diabetes. The disruption of blood flow may also occur more acutely. This includes but is not limited to thrombus formation that results in reduction of blood flow through the coronary arteries, lodging of an embolism in a cerebral ischemia and similar events in the kidney and limbs.
For either a gradual or an acute time course, the earlier the ischemic/hypoxic condition can be detected and the sooner palliative therapy can be applied, the better will be the outcome for tissue and organism. For example, early detection of the diabetic-mediated ischemia in extremities might avoid amputation and early detection and relief of acute blockages to the heart or brain significantly reduces mortality and morbidity.
Unfortunately, early detection of ischemia/hypoxia is often not possible. For example, the ECG which is the primary early diagnostic tool for acute coronary syndromes is less than 40% sensitive. For stroke patients the only tools available are either a CT scan or MRI, both of which can only determine if the patient has a hemorrhage in the first several hours after symptoms began. Only much later, when it is too late to administer the only treatment of ischemic stroke, thrombolytic therapy, are these imaging methods able to determine if an ischemic stroke has occurred. Thrombolytic therapy must be administered within 3 hours of symptoms. For other organs there are no well established methods for early detection of ischemia. Thus a sensitive, accurate and rapid test for ischemia is needed for the diagnosis and treatment of patients.
In the final stages of ischemia, when cells begin to die (necrosis), they may release some of their contents into the blood. These are primarily intracellular proteins that are released because the normal barrier to containment, the cell's membrane, is compromised by biochemical changes associated with death. Often these molecules are tissue specific, for example, cardiac troponin in the case of the heart. Although these molecules accurately reflect the presence of disease, they generally require several hours after symptoms occur to reach levels of significance in blood and they are only released from dead or dying cells. Thus in addition to sensitivity and accuracy an important feature of a test for ischemia would be its ability to detect the ischemic state well before necrosis.
In addition to the release of molecules from the ischemic tissue that are markers of necrosis, the ischemic event may generate a series of biochemical changes that can result in the change in concentration of molecules within the blood or other fluids that do not originate from the ischemic tissue. These are referred to in this invention as ischemic markers. The generation of ischemic markers can occur, for example, when the ischemic tissue generates molecules that are then converted to different molecules by a non-ischemic tissue or the ischemic tissue generates molecules that activate, from non-ischemic tissue, the release of different molecules into body fluids. Examples of this are the generation of norepinephrine, TNFα and natriuretic peptides by ischemic cardiac or cerebral tissue. The norepinephrine, TNFα and natriuretic peptides so generated activate lipolysis in adipose tissue resulting in the elevation of free fatty acids in blood. In another example, sphingosine released from the ischemic myocardium is converted to sphingosine-1-phosphate in platelets and may then be detected in blood (Yatomi, et al. (1997) Journ. Biol. Chem. vol. 272: pages 5291-5297; U.S. Pat. No. 6,210,976).
In addition to diagnosing the presence of disease, levels of ischemic markers may predict risk of future deleterious events. Numerous in vitro studies reveal that elevated free fatty acids (FFA) are potent perturbants of many cellular functions, and clinical evidence for a direct role of FFA is strongly suggested by a number of studies pointing to their potential role in cardiovascular disease (Leaf A. Circulation 104: 744-745, 2001; Oliver M F and Opie L H. The Lancet 343: 155-158, 1994; Paolisso G, et al. American Journal of Cardiology 80: 932-937, 1997; Carlsson M, et al. Arterioscler throm Vasc Biol 20: 1588-1594, 2000). Elevations in total plasma FFA are associated with increased risks of cardiac arrhythmias (Kurien V A and Oliver M F. Br Heart J 32: 556, 1970; Kurien V A and Oliver M F. The Lancet April 18: 813-815, 1970; Leaf A. Circulation 104: 744-745, 2001; Oliver M F. Am J Med 112: 305-311, 2002) and death in non-acute cardiovascular disease (Jouven X, et al. Circulation 104: 756-761, 2001; Pilz S, et al. J Clin Endocrinol Metab 91: 2542-2547, 2006) and therefore are a particularly unwelcome consequence of acute coronary syndrome (ACS).
Moreover, these molecules, at sufficient levels, may themselves mediate cellular effects that result in deleterious outcomes. For example, a large long term study of apparently healthy men revealed that increasing levels of total serum free fatty acids (FFA), although within the normal range, were associated with an increased risk of sudden death 22 years later. It was speculated that this increased rate of death was a consequence of FFA induced cardiac arrhythmias. In another example, use of the combination of glucose-insulin-potassium (GIK) in patients suffering from acute myocardial infarcts, produced a significant reduction in mortality relative to patients who did not receive GIK. One theory for this beneficial effect is the reduction in serum total FFA produced by GIK. Thus there is a need to be able to monitor ischemic markers to evaluate longer term risk of disease and to help to decide on the type of therapeutic intervention to reduce the increased risk associated with what may be a chronic ischemic state.